Sign up to receive free email alerts when patent applications with chosen keywords are publishedSIGN UP

Abstract:

This disclosure concerns novel compounds of Formula (I) as defined in the
specification and compositions comprising such novel compounds. These
compounds are useful antiviral agents, especially in inhibiting the
function of the NS5A protein encoded by Hepatitis C virus (HCV). Thus,
the disclosure also concerns a method of treating HCV related diseases or
conditions by use of these novel compounds or a composition comprising
such novel compounds.
##STR00001##

Claims:

2. A composition comprising a compound of claim 1, or a pharmaceutically
acceptable salt thereof, and a pharmaceutically acceptable carrier.

3. A method of treating an HCV infection in a patient, comprising
administering to the patient a therapeutically effective amount of a
compound of claim 1, or a pharmaceutically acceptable salt thereof.

[0002] The present disclosure is generally directed to antiviral
compounds, and more specifically directed to compounds which can inhibit
the function of the NS5A protein encoded by Hepatitis C virus (HCV),
compositions comprising such compounds, and methods for inhibiting the
function of the NS5A protein.

BACKGROUND OF THE DISCLOSURE

[0003] HCV is a major human pathogen, infecting an estimated 170 million
persons worldwide--roughly five times the number infected by human
immunodeficiency virus type 1. A substantial fraction of these HCV
infected individuals develop serious progressive liver disease, including
cirrhosis and hepatocellular carcinoma.

[0004] The current standard of care for HCV, which employs a combination
of pegylated-interferon and ribavirin, has a non-optimal success rate in
achieving sustained viral response and causes numerous side effects.
Thus, there is a clear and long-felt need to develop effective therapies
to address this undermet medical need.

[0005] HCV is a positive-stranded RNA virus. Based on a comparison of the
deduced amino acid sequence and the extensive similarity in the 5'
untranslated region, HCV has been classified as a separate genus in the
Flaviviridae family. All members of the Flaviviridae family have
enveloped virions that contain a positive stranded RNA genome encoding
all known virus-specific proteins via translation of a single,
uninterrupted, open reading frame.

[0006] Considerable heterogeneity is found within the nucleotide and
encoded amino acid sequence throughout the HCV genome due to the high
error rate of the encoded RNA dependent RNA polymerase which lacks a
proof-reading capability. At least six major genotypes have been
characterized, and more than 50 subtypes have been described with
distribution worldwide. The clinical significance of the genetic
heterogeneity of HCV has demonstrated a propensity for mutations to arise
during monotherapy treatment, thus additional treatment options for use
are desired. The possible modulator effect of genotypes on pathogenesis
and therapy remains elusive.

[0007] The single strand HCV RNA genome is approximately 9500 nucleotides
in length and has a single open reading frame (ORF) encoding a single
large polyprotein of about 3000 amino acids. In infected cells, this
polyprotein is cleaved at multiple sites by cellular and viral proteases
to produce the structural and non-structural (NS) proteins. In the case
of HCV, the generation of mature non-structural proteins (NS2, NS3, NS4A,
NS4B, NS5A, and NS5B) is effected by two viral proteases. The first one
is believed to be a metalloprotease and cleaves at the NS2-NS3 junction;
the second one is a serine protease contained within the N-terminal
region of NS3 (also referred to herein as NS3 protease) and mediates all
the subsequent cleavages downstream of NS3, both in cis, at the NS3-NS4A
cleavage site, and in trans, for the remaining NS4A-NS4B, NS4B-NS5A,
NS5A-NS5B sites. The NS4A protein appears to serve multiple functions by
both acting as a cofactor for the NS3 protease and assisting in the
membrane localization of NS3 and other viral replicase components. The
formation of a NS3-NS4A complex is necessary for proper protease activity
resulting in increased proteolytic efficiency of the cleavage events. The
NS3 protein also exhibits nucleoside triphosphatase and RNA helicase
activities. NS5B (also referred to herein as HCV polymerase) is a
RNA-dependent RNA polymerase that is involved in the replication of HCV
with other HCV proteins, including NS5A, in a replicase complex.

[0012] X and Z, together with the carbon atoms to which they are attached,
form a five- to eight-membered aromatic or non-aromatic fused ring
optionally containing one or two heteroatoms independently selected from
nitrogen, oxygen, and sulfur; wherein the five- to eight-membered ring is
optionally substituted with one, two, or three substitutents
independently selected from alkoxy, alkoxyalkyl, alkoxycarbonyl, alkyl,
alkylsulfonyl, aryl, arylalkyl, arylsulfonyl, carboxy, formyl, halo,
haloalkoxy, haloalkyl, hydroxy, hydroxyalkyl, --NRaRb,
(NRaRb)alkyl, (NRaRb)carbonyl, oxo, and spirocycle;

[0013] X' is hydrogen (H) or halogen and Z' is hydrogen; or

[0014] X' and Z', together with the carbon atoms to which they are
attached, form a five- to eight-membered aromatic or non-aromatic fused
ring optionally containing one or two heteroatoms independently selected
from nitrogen, oxygen, and sulfur; wherein the five- to eight-membered
ring is optionally substituted with one, two, or three substitutents
independently selected from alkoxy, alkoxyalkyl, alkoxycarbonyl, alkyl,
alkylsulfonyl, aryl, arylalkyl, arylsulfonyl, carboxy, formyl, halo,
haloalkoxy, haloalkyl, hydroxy, hydroxyalkyl, --NRaRb,
(NRaRb)alkyl, (NRaRb)carbonyl, oxo, and spirocycle;

[0015] Y and Y' are each independently --CH2--, --CH2CH2--,
or --CH2O--, wherein the --CH2O-- is drawn such that the oxygen
atom is bound to the carbon atom substituted with Rv and Rq or
Rv' and Rq';

[0016] Rp is hydrogen or C1 to C4 alkyl;

[0017] Rq is hydrogen, alkyl, or halo; or

[0018] Rp and Rq, together with the carbon atoms to which they
are attached, form a cycloalkyl ring;

[0019] Rv is selected from hydrogen, alkyl, halo, and hydroxy; or

[0020] Rv and Rq, together with the carbon atom to which they
are attached, form an ethylenyl group or a cycloalkyl ring;

[0021] Rp' is hydrogen or C1 to C4 alkyl;

[0022] Rq' is hydrogen, alkyl, or halo; or

[0023] Rp' and Rq', together with the carbon atoms to which they
are attached, form a cycloalkyl ring;

[0024] Rv' are independently selected from hydrogen, alkyl, halo, and
hydroxy; or

[0025] Rv' and Rq', together with the carbon atom to which they
are attached, form an ethylenyl group or a cycloalkyl ring;

[0026] Rw and Rw' are independently selected from hydrogen and
alkyl;

[0027] R1 is hydrogen or --C(O)Rx;

[0028] R2 is hydrogen or --C(O)Ry;

[0029] Rx and Ry are independently selected from cycloalkyl,
heteroaryl, heterocyclyl, alkoxy, and alkyl, said alkyl being substituted
by one or more substituents independently selected from aryl, alkenyl,
cycloalkyl, heterocyclyl, heteroaryl, --OR3, --C(O)OR4,
--NRaRb, and --C(O)NRcRd,

[0030] wherein any said aryl and heteroaryl may optionally be substituted
with one or more substituents independently selected from alkenyl, alkyl,
haloalkyl, arylalkyl, heterocyclyl, heterocyclylalkyl, halogen, cyano,
nitro, --C(O)OR4, --OR5, --NRaRb,
(NRaRb)alkyl, and (MeO)(HO)P(O)O--, and

[0031] wherein any said cycloalkyl and heterocyclyl may optionally be
fused onto an aromatic ring and may optionally be substituted with one or
more substituents independently selected from alkyl, hydroxyl, halogen,
aryl, --NRaRb, oxo, and --C(O)OR4;

[0032] R3 is hydrogen, alkyl, or arylalkyl;

[0033] R4 is alkyl or arylalkyl;

[0034] R5 is hydrogen, alkyl, or arylalkyl;

[0035] Ra and Rb are independently selected from hydrogen,
alkyl, cycloalkyl, arylalkyl, heteroaryl, --C(O)R6, --C(O)OR7,
--C(O)NRcRd, and (NRcRd)alkyl, or alternatively,
Ra and Rb, together with the nitrogen atom to which they are
attached, form a five- or six-membered ring or bridged bicyclic ring
structure, wherein said five- or six-membered ring or bridged bicyclic
ring structure optionally may contain one or two additional heteroatoms
independently selected from nitrogen, oxygen, and sulfur and may contain
one, two, or three substituents independently selected from C1 to
C6 alkyl, C1 to C4 haloalkyl, aryl, hydroxyl, C1 to
C6 alkoxy, C1 to C4 haloalkoxy, and halogen;

[0036] R6 is alkyl;

[0037] R7 is alkyl, arylalkyl, cycloalkyl, or haloalkyl; and

[0038] Rc and Rd are independently selected from hydrogen,
alkyl, arylalkyl, and cycloalkyl.

[0039] In a first embodiment of the first aspect the present disclosure
provides a compound of Formula (I) further characterized by Formula (Ia):

##STR00004##

or a pharmaceutically acceptable salt or a tautomer thereof, wherein:

[0040] X is hydrogen or chloro (Cl) and Z is hydrogen; or

[0041] X and Z, together with the carbon atoms to which they are attached,
form a six-membered aromatic or non-aromatic fused ring;

[0042] X' is hydrogen or chloro (Cl) and Z' is hydrogen; or

[0043] X' and Z', together with the carbon atoms to which they are
attached, form a six-membered aromatic or non-aromatic fused ring;

[0044] Y is --CH2--, --CH2CH2--, or --CH2O--, wherein
the --CH2O-- is drawn such that the oxygen atom is bound to the
carbon atom substituted with Rv and Rq;

[0045] Rp is hydrogen or C1 to C4 alkyl;

[0046] Rq is hydrogen, alkyl, or halo; or

[0047] Rp and Rq, together with the carbon atoms to which they
are attached, form a cycloalkyll ring; and

[0048] Rv is selected from hydrogen, alkyl, halo, and hydroxy; or

[0049] Rv and Rq, together with the carbon atom to which they
are attached, form an ethylenyl group or a cycloalkyl ring.

[0050] In a second embodiment of the first aspect the present disclosure
provides a compound of Formula (I) further characterized by Formula (Ib):

##STR00005##

or a pharmaceutically acceptable salt or a tautomer thereof.

[0051] In a third embodiment of the first aspect the present disclosure
provides a compound of Formula (I) further characterized by Formula (Ic):

##STR00006##

or a pharmaceutically acceptable salt or a tautomer thereof.

[0052] In a fourth embodiment of the first aspect the present disclosure
provides a compound of Formula (I) further characterized by Formula (Id):

##STR00007##

or a pharmaceutically acceptable salt or a tautomer thereof.

[0053] In a fifth embodiment of the first aspect the present disclosure
provides a compound of Formula (Ia), or a pharmaceutically acceptable
salt thereof, wherein:

[0054] R1 is --C(O)Rx;

[0055] R2 is --C(O)Ry;

[0056] Rx and Ry are independently alkyl substituted by at least
one --NRaRb, characterized by Formula (A):

##STR00008##

wherein:

[0057] m is 0 or 1;

[0058] R8 is hydrogen or alkyl;

[0059] R9 is selected from hydrogen, cycloalkyl, aryl, heteroaryl,
heterocyclyl, and alkyl optionally substituted with a substituent
selected from aryl, alkenyl, cycloalkyl, heterocyclyl, heteroaryl,
heterobicyclyl, --OR3, --C(O)OR4, --NRaRb, and
--C(O)NRcRd, wherein any said aryl and heteroaryl may
optionally be substituted with one or more substituents independently
selected from alkyl, haloalkyl, arylalkyl, heterocyclyl,
heterocyclylalkyl, halogen, cyano, nitro, --C(O)OR4, --OR5,
--NRaRb, (NRaRb)alkyl, and (MeO)(HO)P(O)O--, and

[0060] wherein any said cycloalkyl and heterocyclyl may optionally be
fused onto an aromatic ring and may optionally be substituted with one or
more substituents independently selected from alkyl, hydroxyl, halogen,
aryl, --NRaRb, oxo, and --C(O)OR4; and

[0061] R3, R4, R5, Ra, Rb, Rc, and Rd
are defined as in Formula (I).

[0062] In a sixth embodiment of the first aspect the present disclosure
provides a compound of Formula (Ia) or a pharmaceutically acceptable salt
thereof, wherein

[0097] In a tenth embodiment of the first aspect the present disclosure
provides a compound of Formula (Ia) or a pharmaceutically acceptable salt
thereof, wherein

[0098] R1 is --C(O)Rx;

[0099] R2 is --C(O)Ry;

[0100] Rx and Ry are cycloalkyl independently selected from:

##STR00012##

wherein

[0101] j is 0, 1, 2, or 3;

[0102] k is 0, 1, or 2;

[0103] n is 0 or an integer selected from 1 through 4;

[0104] each R13 is independently selected from hydrogen, C1 to
C6 alkyl, C1 to C4 haloalkyl, C1 to C6 alkoxy,
halogen, hydroxyl, cyano, and nitro; and

[0105] Ra and Rb are each independently hydrogen, C1 to
C6 alkyl, or --C(O)OR7, wherein R7 is C1 to C6
alkyl.

[0106] In an eleventh embodiment of the first aspect the present
disclosure provides a compound of Formula (Ia) or a pharmaceutically
acceptable salt thereof, wherein

[0107] R1 is --C(O)Rx;

[0108] R2 is --C(O)R'';

[0109] Rx and Ry are independently arylalkyl, wherein aryl part
of said arylalkyl may optionally be substituted with
(NRaRb)alkyl; and

[0110] Ra and Rb are independently hydrogen, C1 to C6
alkyl, or benzyl, or alternatively, Ra and Rb, together with
the nitrogen atom to which they are attached, form a five- or
six-membered ring selected from

##STR00013##

wherein R15 is hydrogen, C1 to C6 alkyl, or benzyl.

[0111] In a twelfth embodiment of the first aspect the present disclosure
provides a compound of Formula (Ia) or a pharmaceutically acceptable salt
thereof, wherein

[0112] R1 and R2 are the same and are selected from the group
consisting of:

##STR00014## ##STR00015##

wherein a squiggle bond () in the structure indicates that a stereogenic
center to which the bond is attached can take either (R)-- or (S)--
configuration so long as chemical bonding principles are not violated.

[0113] In a thirteenth embodiment of the first aspect the present
disclosure provides a compound of Formula (I), or a pharmaceutically
acceptable salt thereof, wherein

[0114] R1 is --C(O)Rx;

[0115] R2 is --C(O)Ry; and

[0116] Rx and Ry are both t-butoxy.

[0117] In a fourteenth embodiment of the first aspect the present
disclosure provides a compound of Formula (I), or a pharmaceutically
acceptable salt thereof, wherein

[0118] R1 and R2 are both hydrogen.

[0119] In a second aspect the present disclosure provides a compound of
Formula (II):

[0122] Rp is hydrogen or C1 to C4 alkyl, and Rq is
hydrogen, or alternatively, Rp and Rq, together with the carbon
atoms to which they are attached, form a cyclopropyl ring;

[0123] Rp' is hydrogen or C1 to C4 alkyl, and Rq' is
hydrogen, or alternatively, Rp' and Rq', together with the
carbon atoms to which they are attached, form a cyclopropyl ring;

[0124] R1 is hydrogen or --C(O)Rx;

[0125] R2 is hydrogen or --C(O)Ry;

[0126] Rx and Ry are independently selected from cycloalkyl,
heteroaryl, heterocyclyl, alkoxy, and alkyl, said alkyl being substituted
by one or more substituents independently selected from aryl, alkenyl,
cycloalkyl, heterocyclyl, heteroaryl, --OR3, --C(O)OR4,
--NRaRb, and --C(O)NRcRd,

[0127] wherein any said aryl and heteroaryl may optionally be substituted
with one or more substituents independently selected from alkyl,
haloalkyl, arylalkyl, heterocyclyl, heterocyclylalkyl, halogen, cyano,
nitro, --C(O)OR4, --OR5, --NRaRb,
(NRaRb)alkyl, and (MeO)(HO)P(O)O--, and

[0128] wherein any said cycloalkyl and heterocyclyl may optionally be
fused onto an aromatic ring and may optionally be substituted with one or
more substituents independently selected from alkyl, hydroxyl, halogen,
aryl, --NRaRb, oxo, and --C(O)OR4;

[0129] R3 is hydrogen, alkyl, or arylalkyl;

[0130] R4 is alkyl or arylalkyl;

[0131] R5 is hydrogen, alkyl, or arylalkyl;

[0132] Ra and Rb are independently selected from hydrogen,
alkyl, cycloalkyl, arylalkyl, heteroaryl, --C(O)R6, --C(O)OR7,
--C(O)NRcRd, and (NRcRd)alkyl, or alternatively,
Ra and Rb, together with the nitrogen atom to which they are
attached, form a five- or six-membered ring or bridged bicyclic ring
structure, wherein said five- or six-membered ring or bridged bicyclic
ring structure optionally may contain one or two additional heteroatoms
independently selected from nitrogen, oxygen, and sulfur and may contain
one, two, or three substituents independently selected from C1 to
C6 alkyl, C1 to C4 haloalkyl, aryl, hydroxyl, C1 to
C6 alkoxy, C1 to C4 haloalkoxy, and halogen;

[0133] R6 is alkyl;

[0134] R7 is alkyl, arylalkyl, or haloalkyl; and

[0135] Rc and Rd are independently selected from hydrogen,
alkyl, arylalkyl, and cycloalkyl.

[0136] In a third aspect the present disclosure provides a composition
comprising a compound of Formula (I), or a pharmaceutically acceptable
salt thereof, and a pharmaceutically acceptable carrier, wherein Formula
(I) is defined according to any of the embodiments described above in the
first aspect of the present disclosure.

[0137] In a first embodiment of the third aspect the composition further
comprises at least one additional compound having anti-HCV activity.

[0138] In a second embodiment of the third aspect at least one of the
additional compounds is an interferon or a ribavirin.

[0139] In a third embodiment of the third aspect the interferon is
selected from interferon alpha 2B, pegylated interferon alpha, consensus
interferon, interferon alpha 2A, and lymphoblastoid interferon tau.

[0140] In a fourth embodiment of the third aspect the present disclosure
provides a composition comprising a compound of Formula (I), or a
pharmaceutically acceptable salt thereof, a pharmaceutically acceptable
carrier, and at least one additional compound having anti-HCV activity,
wherein at least one of the additional compounds is selected from
interleukin 2, interleukin 6, interleukin 12, a compound that enhances
the development of a type 1 helper T cell response, interfering RNA,
anti-sense RNA, Imiquimod, ribavirin, an inosine 5'-monophospate
dehydrogenase inhibitor, amantadine, and rimantadine.

[0141] In a fifth embodiment of the third aspect the present disclosure
provides a composition comprising a compound of Formula (I), or a
pharmaceutically acceptable salt thereof, a pharmaceutically acceptable
carrier, and at least one additional compound having anti-HCV activity,
wherein at least one of the additional compounds is effective to inhibit
the function of a target selected from HCV metalloprotease, HCV serine
protease, HCV polymerase, HCV helicase, HCV NS4B protein, HCV entry, HCV
assembly, HCV egress, HCV NS5A protein, and IMPDH for the treatment of an
HCV infection.

[0142] In a fourth aspect the present disclosure provides a method of
treating an HCV infection in a patient, comprising administering to the
patient a therapeutically effective amount of a compound of Formula (I),
or a pharmaceutically acceptable salt thereof, wherein Formula (I) is
defined according to any of the embodiments described above in the first
aspect of the present disclosure. In a first embodiment of the fourth
aspect the method further comprises administering at least one additional
compound having anti-HCV activity prior to, after or simultaneously with
the compound of Formula (I), or a pharmaceutically acceptable salt
thereof.

[0143] In a second embodiment of the fourth aspect at least one of the
additional compounds is an interferon or a ribavirin.

[0144] In a third embodiment of the fourth aspect the interferon is
selected from interferon alpha 2B, pegylated interferon alpha, consensus
interferon, interferon alpha 2A, and lymphoblastoid interferon tau.

[0145] In a fourth embodiment of the fourth aspect the present disclosure
provides a method of treating an HCV infection in a patient, comprising
administering to the patient a therapeutically effective amount of a
compound of Formula (I), or a pharmaceutically acceptable salt thereof,
and at least one additional compound having anti-HCV activity prior to,
after or simultaneously with the compound of Formula (I), or a
pharmaceutically acceptable salt thereof, wherein at least one of the
additional compounds is selected from interleukin 2, interleukin 6,
interleukin 12, a compound that enhances the development of a type 1
helper T cell response, interfering RNA, anti-sense RNA, Imiquimod,
ribavirin, an inosine 5'-monophospate dehydrogenase inhibitor,
amantadine, and rimantadine.

[0146] In a fifth embodiment of the fourth aspect the present disclosure
provides a method of treating an HCV infection in a patient, comprising
administering to the patient a therapeutically effective amount of a
compound of Formula (I), or a pharmaceutically acceptable salt thereof,
and at least one additional compound having anti-HCV activity prior to,
after or simultaneously with the compound of Formula (I), or a
pharmaceutically acceptable salt thereof, wherein at least one of the
additional compounds is effective to inhibit the function of a target
selected from HCV metalloprotease, HCV serine protease, HCV polymerase,
HCV helicase, HCV NS4B protein, HCV entry, HCV assembly, HCV egress, HCV
NS5A protein, and IMPDH for the treatment of an HCV infection.

[0147] The compounds of the present disclosure can be effective to inhibit
the function of the HCV NS5A protein. In particular, the compounds of the
present disclosure can be effective to inhibit the HCV 1b genotype or
multiple genotypes of HCV. Therefore, this disclosure also encompasses:
(1) compositions comprising a compound of Formula (I), or a
pharmaceutically acceptable salt thereof, and a pharmaceutically
acceptable carrier; and (2) a method of treating an HCV infection in a
patient, comprising administering to the patient a therapeutically
effective amount of a compound of Formula (I), or a pharmaceutically
acceptable salt thereof. Other aspects of the present disclosure may
include suitable combinations of embodiments disclosed herein.

[0148] Yet other aspects and embodiments may be found in the description
provided herein.

[0149] The description of the present disclosure herein should be
construed in congruity with the laws and principals of chemical bonding.
In some instances it may be necessary to remove a hydrogen atom in order
to accommodate a substituent at any given location.

[0150] Certain features of the structure of Formula (I) are further
illustrated below:

##STR00018##

[0151] In Formula (I), as depicted above, the "pyrrolidinyl-imidazole"
moiety on the left side of the "linker" is independent from the
"pyrrolidinyl-imidazole" moiety on the right side of the linker group in
respect to, e.g., (1) tautomer form of imidazole ring, (2) absolute
configuration of the stereogenic centers on the pyrrolidine ring, and (3)
substituents on the pyrrolidine nitrogen, i.e., R1 and R2 are
independent from each other, although in some circumstances they are
preferably the same.

[0152] It should be understood that the depiction of a pyrrolidine moiety
on the "left" side or on the "right" side is for illustration purpose
only, which does not in any way limit the scope of the disclosure.

[0153] In the linker group of Formula (I), the linkage between "L" and the
two benzene rings encompasses all the following combinations:

##STR00019##

wherein the "para-para," "para-meta," and "meta-meta" linkages are
preferred.

[0154] Likewise, in Formula (I), when L is a phenylene

##STR00020##

group, it can link to the adjacent two benzene rings by the following
manners:

##STR00021##

wherein the "para" and "meta" arrangements are preferred, and the "para"
arrangement is the more preferred.

[0155] In Formula (I), when L is a vinylene (--CH═CH--) group, it can
take either trans- or cis-configuration, as depicted below:

##STR00022##

[0156] In Formula (I), when L is a cyclopropylene

##STR00023##

group, the two benzene substituents can be either trans- or cis- to each
other, forming one of the following four configurations:

##STR00024##

[0157] In a pyrrolidine ring of a pyrrolidinyl-imidazole moiety, the
stereogenic carbon center to which the imidazole group is attached can
take either (R)-- or (S)-- configuration as depicted below:

##STR00025##

[0158] When a cyclopropyl ring is fused onto a pyrrolidine ring of a
pyrrolidinyl-imidazole moiety, i.e., when (Rp, Rq) together is
--CH2--, the CH2 group of the fused cyclopropyl ring can take
either α- or β-position relative to the pyrrolidine ring, as
depicted below:

##STR00026##

[0159] Thus, this disclosure is intended to cover all possible
stereoisomers even when a single stereoisomer, or no stereochemistry, is
described in a structure.

[0160] In Formula (I), the linkage between a benzene ring of the linker
group and an imidazole ring of a pyrrolidinyl-imidazole moiety can take
place in either the C-4 or the C-5 position (see below) of the imidazole
ring. As a person of ordinary skill in the art would understand, due to
tautomerization of the imidazole ring, a bonding of a benzene ring to the
C-4 position may be equivalent to a bonding of the benzene ring to the
C-5 position, as shown in the following equation:

##STR00027##

[0161] The sample principle also applies to substituent X or X'.

[0162] Thus, this disclosure is intended to cover all possible tautomers
even when a structure depicts only one of them.

[0163] In this disclosure, a floating bond

##STR00028##

or a floating substituent (e.g., --R13) on a structure indicates
that the bond or substituent can attach to any available position of the
structure by removal of a hydrogen from the available position. It should
be understood that in a bicyclic or polycyclic ring structure, unless
specifically defined otherwise, the position of a floating bond or a
floating substituent does not limit the position of such bond or
substituent to a specific ring. Thus, the following two substituents
should be construed to be equivalent:

##STR00029##

[0164] It should be understood that the compounds encompassed by the
present disclosure are those that are suitably stable for use as
pharmaceutical agent.

[0165] It is intended that the definition of any substituent or variable
at a particular location in a molecule be independent of its definitions
elsewhere in that molecule. For example, for substituent
(R10)n, when n is 2, each of the two R10 groups may be the
same or different.

[0166] All patents, patent applications, and literature references cited
in the specification are herein incorporated by reference in their
entirety. In the case of inconsistencies, the present disclosure,
including definitions, will prevail.

DEFINITIONS

[0167] Definitions have been provided above for each of the groups
defined. In addition, the following definitions shall be used.

[0168] As used herein, the singular forms "a", "an", and "the" include
plural reference unless the context clearly dictates otherwise.

[0169] Unless stated otherwise, all aryl, cycloalkyl, heteroaryl, and
heterocyclyl groups of the present disclosure may be substituted as
described in each of their respective definitions. For example, the aryl
part of an arylalkyl group may be substituted as described in the
definition of the term "aryl."

[0170] The term "acetyl," as used herein, refers to --C(O)CH3.

[0171] The term "alkenyl," as used herein, refers to a monovalent,
straight or branched hydrocarbon chain having one or more, preferably one
to two, double bonds therein. The double bond of an alkenyl group can be
unconjugated or conjugated to another unsaturated group. Suitable alkenyl
groups include, but are not limited to, C2 to C10 alkenyl
groups, such as vinyl, allyl, butenyl, pentenyl, hexenyl, butadienyl,
pentadienyl, hexadienyl, 2-ethylhexenyl, 2-propyl-2-butenyl,
4-(2-methyl-3-butene)-pentenyl. An alkenyl group can be unsubstituted or
substituted with one or two suitable substituents.

[0172] The term "alkoxy," as used herein, refers to an alkyl group
attached to the parent molecular moiety through an oxygen atom.
Representative examples of alkoxy group include, but are not limited to,
methoxy (CH3O--), ethoxy (CH3CH2O--), and t-butoxy
((CH3)3CO--).

[0173] The term "alkoxyalkyl," as used herein, refers to an alkyl group
substituted with one, two, or three alkoxy groups.

[0174] The term "alkoxycarbonyl," as used herein, refers to an alkoxy
group attached to the parent molecular moiety through a carbonyl group.

[0175] The term "alkyl," as used herein, refers to a group derived from a
straight or branched chain saturated hydrocarbon by removal of a hydrogen
from one of the saturated carbons. The alkyl group preferably contains
from one to ten carbon atoms.

[0176] Representative examples of alkyl group include, but are not limited
to, methyl, ethyl, isopropyl, and tert-butyl.

[0177] The term "alkylcarbonyl," as used herein, refers to an alkyl group
attached to the parent molecular moiety through a carbonyl group.
Representative examples of alkylcarbonyl group include, but are not
limited to, acetyl (--C(O)CH3), propanoyl (--C(O)CH2CH3),
n-butyryl (--C(O)CH2CH2CH3), and 2,2-dimethylpropanoyl or
pivaloyl (--C(O)C(CH3)3).

[0178] The term "alkylsulfonyl," as used herein, refers to an alkyl group
attached to the parent molecular moiety through a sulfonyl group.

[0179] The term "allyl," as used herein, refers to the
--CH2CH═CH2 group.

[0180] The term "aryl," as used herein, refers to a group derived from an
aromatic carbocycle by removal of a hydrogen atom from an aromatic ring.
The aryl group can be monocyclic, bicyclic or polycyclic, wherein in
bicyclic or polycyclic aryl group, the aromatic carbocycle can be fused
onto another four- to six-membered aromatic or non-aromatic carbocycle.
Representative examples of aryl groups include, but are not limited to,
phenyl, indanyl, indenyl, naphthyl, and 1,2,3,4-tetrahydronaphth-5-yl.

The term "arylalkyl," as used herein, refers to an alkyl group
substituted with one, two, or three aryl groups, wherein aryl part of the
arylalkyl group may optionally be substituted by one to five substituents
independently selected from C1 to C6 alkyl, C1 to C4
haloalkyl, C1 to C6 alkoxy, halogen, cyano, and nitro groups.
Represented examples of arylalkyl include, but are not limited to,
benzyl, 2-phenyl-1-ethyl (PhCH2CH2--), (naphth-1-yl)methyl, and
(naphth-2-yl)methyl.

[0181] The term "arylsulfonyl," as used herein, refers to an aryl group
attached to the parent molecular moiety through a sulfonyl group.

[0182] The term "benzyl," as used herein, refers to a methyl group on
which one of the hydrogen atoms is replaced by a phenyl group, wherein
said phenyl group may optionally be substituted by one to five
substituents independently selected from methyl, trifluoromethyl
(--CF3), methoxy (--OCH3), halogen, and nitro (--NO2).

[0183] Representative examples of benzyl group include, but are not
limited to, PhCH2--, 4-MeO--C6H4CH2--, and
2,4,6-tri-methyl-C6H4CH2--.

[0184] The term "bridged bicyclic ring," as used herein, refers to a ring
structure comprising a bridgehead between two of the ring members,
wherein the ring and the bridgehead optionally may independently comprise
one or more, preferably one to two, heteroatoms independently selected
from nitrogen, oxygen, and sulfur. Illustrated examples of a bridged
bicyclic ring structure include, but are not limited to:

##STR00030##

[0185] The terms "Cap" and "cap," as used herein, refer to the group which
is placed on the nitrogen atom of the pyrrolidine ring in the compounds
of formula (I). It should be understood that "Cap" or "cap" can also
refer to the reagent which is a precursor to the final "cap" in compounds
of formula (I) and is used as one of the starting materials in the
reaction to append a group on the pyrrolidine nitrogen that results in
the final product, a compound which contains the functionalized
pyrrolidine that will be present in the compound of formula (I).

[0186] The term "carbonyl," as used herein, refers to --C(O)--.

[0187] The term "carboxyl," or "carboxy," as used herein, refers to
--CO2H.

[0188] The term "cyano," as used herein, refers to --CN.

[0189] The term "cycloalkyl," as used herein, refers to a group derived
from a saturated carbocycle, having preferably three to eight carbon
atoms, by removal of a hydrogen atom from the saturated carbocycle,
wherein the saturated carbocycle can optionally be fused onto one or two
other aromatic or nonaromatic carbocycles. Representative examples of
cycloalkyl groups include, but are not limited to, cyclopropyl,
cyclopentyl, cyclohexyl, and 1,2,3,4-tetrahydronaphth-1-yl.

[0190] The term "formyl," as used herein, refers to --CHO.

[0191] The terms "halo" and "halogen," as used herein, refer to F, Cl, Br,
or I. The term "haloalkoxy," as used herein, refers to a haloalkyl group
attached to the parent molecular moiety through an oxygen atom.

[0192] The term "haloalkyl," as used herein, refers to an alkyl group
substituted by at least one halogen atom. The haloalkyl group can be an
alkyl group of which all hydrogen atoms are substituted by halogens.
Representative examples of haloalkyl include, but are not limited to,
trifluoromethyl (CF3--), 1-chloroethyl (ClCH2CH2--), and
2,2,2-trifluoroethyl (CF3CH2--).

[0193] The term "heteroaryl," as used herein, refers to group derived from
a monocyclic, bicyclic, or polycyclic compound comprising at least one
aromatic ring comprising one or more, preferably one to three,
heteroatoms independently selected from nitrogen, oxygen, and sulfur, by
removal of a hydrogen atom from an aromatic ring thereof. As is well
known to those skilled in the art, heteroaryl rings have less aromatic
character than their all-carbon counterparts. Thus, for the purposes of
the disclosure, a heteroaryl group need only have some degree of aromatic
character. Illustrative examples of heteroaryl groups include, but are
not limited to, pyridyl, pyridazinyl, pyrimidyl, pyrazyl, triazinyl,
pyrrolyl, pyrazolyl, imidazolyl, (1,2,3,)- and (1,2,4)-triazolyl,
pyrazinyl, pyrimidinyl, tetrazolyl, furyl, thienyl, isoxazolyl,
thiazolyl, isoxazolyl, oxazolyl, indolyl, quinolinyl, isoquinolinyl,
benzisoxazolyl, benzothiazolyl, benzothienyl, and pyrrolopyridinyl.

[0194] The term "heteroarylalkyl," as used herein, refers to an alkyl
group substituted with one, two, or three heteroaryl groups.

[0195] The term "heterobicyclyl," as used herein, refers to a ring
structure comprising two fused or bridged rings that include carbon and
one or more, preferably one to three, heteroatoms independently selected
from nitrogen, oxygen, and sulfur. The heterobicyclic ring structure is a
subset of heterocyclic ring and can be saturated or unsaturated. Examples
of heterobicyclic ring structures include, but are not limited to,
tropane, quinuclidine, and 7-azabicyclo[2.2.1]heptane.

[0196] The term "heterocyclyl," as used herein, refers to a group derived
from a monocyclic, bicyclic, or polycyclic compound comprising at least
one nonaromatic ring comprising one or more, preferably one to three,
heteroatoms independently selected from nitrogen, oxygen, and sulfur, by
removal of a hydrogen atom from the nonaromatic ring. The heterocyclyl
group encompasses the heterobicyclyl group. The heterocyclyl groups of
the present disclosure can be attached to the parent molecular moiety
through a carbon atom or a nitrogen atom in the group. Examples of
heterocyclyl groups include, but are not limited to, morpholinyl,
oxazolidinyl, piperazinyl, piperidinyl, pyrrolidinyl, tetrahydrofuryl,
thiomorpholinyl, and indolinyl.

[0197] The term "heterocyclylalkyl," as used herein, refers to an alkyl
group substituted with one, two, or three heterocyclyl groups.

[0198] The terms "hydroxy" or "hydroxyl," as used herein, refer to --OH.

[0199] The term "hydroxyalkyl," as used herein, refers to an alkyl group
substituted with one, two, or three hydroxy groups.

[0200] The term "nitro," as used herein, refers to --NO2.

[0201] The term "--NRaRb," as used herein, refers to two groups,
Ra and Rb, which are attached to the parent molecular moiety
through a nitrogen atom, or alternatively Ra and Rb, together
with the nitrogen atom to which they are attached, form a 5- or
6-membered ring or a fused- or bridged-bicyclic ring structure optionally
containing one, two, or three additional heteroatom independently
selected from nitrogen, oxygen, and sulfur. The term "--NRcRd"
is defined similarly.

[0202] The term "(NRaRb)alkyl," as used herein, refers to an
alkyl group substituted with one, two, or three --NRaRb groups.
The term "(NRcRd)alkyl" is defined similarly.

[0203] The term "oxo," as used herein, refers to ═O.

[0204] The term "sulfonyl," as used herein, refers to --SO2--.

[0205] The term "trialkylsilyl," as used herein, refers to --SiR3,
wherein each R is C1 to C4 alkyl or phenyl. The three R groups
may be the same or different. Representative examples of "trialkylsilyl"
include, but are not limited to, trimethylsilyl (TMS),
tert-butyldiphenylsilyl (TBDPS), tert-butyldimethylsilyl (TBS or TBDMS),
and triisopropylsilyl (TIPS).

[0206] Asymmetric centers exist in the compounds of the present
disclosure. These centers are designated by the symbols "R" or "S",
depending on the configuration of substituents around the chiral carbon
atom. It should be understood that the disclosure encompasses all
stereochemical isomeric forms, or mixtures thereof, which possess the
ability to inhibit NS5A. Individual stereoisomers of compounds can be
prepared synthetically from commercially available starting materials
which contain chiral centers or by preparation of mixtures of
enantiomeric products followed by separation such as conversion to a
mixture of diastereomers followed by separation or recrystallization,
chromatographic techniques, or direct separation of enantiomers on chiral
chromatographic columns. Starting compounds of particular stereochemistry
are either commercially available or can be made and resolved by
techniques known in the art.

[0207] Certain compounds of the present disclosure may also exist in
different stable conformational forms which may be separable. Torsional
asymmetry due to restricted rotation about an asymmetric single bond, for
example because of steric hindrance or ring strain, may permit separation
of different conformers. The present disclosure includes each
conformational isomer of these compounds and mixtures thereof.

[0208] The term "compounds of the present disclosure", and equivalent
expressions, are meant to embrace compounds of Formula (I), and
pharmaceutically acceptable enantiomers, diastereomers, and salts
thereof. Similarly, references to intermediates are meant to embrace
their salts where the context so permits.

[0209] The present disclosure is intended to include all isotopes of atoms
occurring in the present compounds. Isotopes include those atoms having
the same atomic number but different mass numbers. By way of general
example and without limitation, isotopes of hydrogen include deuterium
and tritium. Isotopes of carbon include 13C and 14C.
Isotopically-labeled compounds of the invention can generally be prepared
by conventional techniques known to those skilled in the art or by
processes analogous to those described herein, using an appropriate
isotopically-labeled reagent in place of the non-labeled reagent
otherwise employed. Such compounds may have a variety of potential uses,
for example as standards and reagents in determining biological activity.
In the case of stable isotopes, such compounds may have the potential to
favorably modify biological, pharmacological, or pharmacokinetic
properties.

[0210] The compounds of the present disclosure can exist as
pharmaceutically acceptable salts. The term "pharmaceutically acceptable
salt," as used herein, represents salts or zwitterionic forms of the
compounds of the present disclosure which are water or oil-soluble or
dispersible, which are, within the scope of sound medical judgment,
suitable for use in contact with the tissues of patients without
excessive toxicity, irritation, allergic response, or other problem or
complication commensurate with a reasonable benefit/risk ratio, and are
effective for their intended use. The salts can be prepared during the
final isolation and purification of the compounds or separately by
reacting a suitable nitrogen atom with a suitable acid. Representative
acid addition salts include acetate, adipate, alginate, citrate,
aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate,
camphorsulfonate; digluconate, glycerophosphate, hemisulfate, heptanoate,
hexanoate, formate, fumarate, hydrochloride, hydrobromide, hydroiodide,
2-hydroxyethanesulfonate, lactate, maleate, mesitylenesulfonate,
methanesulfonate, naphthylenesulfonate, nicotinate,
2-naphthalenesulfonate, oxalate, palmoate, pectinate, persulfate,
3-phenylproprionate, picrate, pivalate, propionate, succinate, tartrate,
trichloroacetate, trifluoroacetate, phosphate, glutamate, bicarbonate,
para-toluenesulfonate, and undecanoate. Examples of acids which can be
employed to form pharmaceutically acceptable addition salts include
inorganic acids such as hydrochloric, hydrobromic, sulfuric, and
phosphoric, and organic acids such as oxalic, maleic, succinic, and
citric.

[0211] Basic addition salts can be prepared during the final isolation and
purification of the compounds by reacting a carboxy group with a suitable
base such as the hydroxide, carbonate, or bicarbonate of a metal cation
or with ammonia or an organic primary, secondary, or tertiary amine. The
cations of pharmaceutically acceptable salts include lithium, sodium,
potassium, calcium, magnesium, and aluminum, as well as nontoxic
quaternary amine cations such as ammonium, tetramethylammonium,
tetraethylammonium, methylamine, dimethylamine, trimethylamine,
triethylamine, diethylamine, ethylamine, tributylamine, pyridine,
N,N-dimethylaniline, N-methylpiperidine, N-methylmorpholine,
dicyclohexylamine, procaine, dibenzylamine, N,N-dibenzylphenethylamine,
and N,N'-dibenzylethylenediamine Other representative organic amines
useful for the formation of base addition salts include ethylenediamine,
ethanolamine, diethanolamine, piperidine, and piperazine.

[0212] When it is possible that, for use in therapy, therapeutically
effective amounts of a compound of Formula (I), as well as
pharmaceutically acceptable salts thereof, may be administered as the raw
chemical, it is possible to present the active ingredient as a
pharmaceutical composition. Accordingly, the disclosure further provides
pharmaceutical compositions, which include therapeutically effective
amounts of compounds of Formula (I) or pharmaceutically acceptable salts
thereof, and one or more, preferably one to three, pharmaceutically
acceptable carriers, diluents, or excipients. The term "therapeutically
effective amount," as used herein, refers to the total amount of each
active component that is sufficient to show a meaningful patient benefit,
e.g., a sustained reduction in viral load. When applied to an individual
active ingredient, administered alone, the term refers to that ingredient
alone. When applied to a combination, the term refers to combined amounts
of the active ingredients that result in the therapeutic effect, whether
administered in combination, serially, or simultaneously. The compounds
of Formula (I) and pharmaceutically acceptable salts thereof, are as
described above. The carrier(s), diluent(s), or excipient(s) must be
acceptable in the sense of being compatible with the other ingredients of
the formulation and not deleterious to the recipient thereof. In
accordance with another aspect of the present disclosure there is also
provided a process for the preparation of a pharmaceutical formulation
including admixing a compound of Formula (I), or a pharmaceutically
acceptable salt thereof, with one or more, preferably one to three,
pharmaceutically acceptable carriers, diluents, or excipients. The term
"pharmaceutically acceptable," as used herein, refers to those compounds,
materials, compositions, and/or dosage forms which are, within the scope
of sound medical judgment, suitable for use in contact with the tissues
of patients without excessive toxicity, irritation, allergic response, or
other problem or complication commensurate with a reasonable benefit/risk
ratio, and are effective for their intended use.

[0213] Pharmaceutical formulations may be presented in unit dose forms
containing a predetermined amount of active ingredient per unit dose.
Dosage levels of between about 0.01 and about 250 milligram per kilogram
("mg/kg") body weight per day, preferably between about 0.05 and about
100 mg/kg body weight per day of the compounds of the present disclosure
are typical in a monotherapy for the prevention and treatment of HCV
mediated disease. Typically, the pharmaceutical compositions of this
disclosure will be administered from about 1 to about 5 times per day or
alternatively, as a continuous infusion. Such administration can be used
as a chronic or acute therapy. The amount of active ingredient that may
be combined with the carrier materials to produce a single dosage form
will vary depending on the condition being treated, the severity of the
condition, the time of administration, the route of administration, the
rate of excretion of the compound employed, the duration of treatment,
and the age, gender, weight, and condition of the patient. Preferred unit
dosage formulations are those containing a daily dose or sub-dose, as
herein above recited, or an appropriate fraction thereof, of an active
ingredient. Generally, treatment is initiated with small dosages
substantially less than the optimum dose of the compound. Thereafter, the
dosage is increased by small increments until the optimum effect under
the circumstances is reached. In general, the compound is most desirably
administered at a concentration level that will generally afford
antivirally effective results without causing any harmful or deleterious
side effects.

[0214] When the compositions of this disclosure comprise a combination of
a compound of the present disclosure and one or more additional
therapeutic or prophylactic agent, both the compound and the additional
agent are usually present at dosage levels of between about 10 to 150%,
and more preferably between about 10 and 80% of the dosage normally
administered in a monotherapy regimen.

[0215] Pharmaceutical formulations may be adapted for administration by
any appropriate route, for example by the oral (including buccal or
sublingual), rectal, nasal, topical (including buccal, sublingual, or
transdermal), vaginal, or parenteral (including subcutaneous,
intracutaneous, intramuscular, intra-articular, intrasynovial,
intrasternal, intrathecal, intralesional, intravenous, or intradermal
injections or infusions) route. Such formulations may be prepared by any
method known in the art of pharmacy, for example by bringing into
association the active ingredient with the carrier(s) or excipient(s).
Oral administration or administration by injection are preferred.

[0216] Pharmaceutical formulations adapted for oral administration may be
presented as discrete units such as capsules or tablets; powders or
granules; solutions or suspensions in aqueous or non-aqueous liquids;
edible foams or whips; or oil-in-water liquid emulsions or water-in-oil
emulsions.

[0217] For instance, for oral administration in the form of a tablet or
capsule, the active drug component can be combined with an oral,
non-toxic pharmaceutically acceptable inert carrier such as ethanol,
glycerol, water, and the like. Powders are prepared by comminuting the
compound to a suitable fine size and mixing with a similarly comminuted
pharmaceutical carrier such as an edible carbohydrate, as, for example,
starch or mannitol. Flavoring, preservative, dispersing, and coloring
agent can also be present.

[0218] Capsules are made by preparing a powder mixture, as described
above, and filling formed gelatin sheaths. Glidants and lubricants such
as colloidal silica, talc, magnesium stearate, calcium stearate, or solid
polyethylene glycol can be added to the powder mixture before the filling
operation. A disintegrating or solubilizing agent such as agar-agar,
calcium carbonate, or sodium carbonate can also be added to improve the
availability of the medicament when the capsule is ingested.

[0219] Moreover, when desired or necessary, suitable binders, lubricants,
disintegrating agents, and coloring agents can also be incorporated into
the mixture. Suitable binders include starch, gelatin, natural sugars
such as glucose or beta-lactose, corn sweeteners, natural and synthetic
gums such as acacia, tragacanth or sodium alginate,
carboxymethylcellulose, polyethylene glycol, and the like. Lubricants
used in these dosage forms include sodium oleate, sodium chloride, and
the like. Disintegrators include, without limitation, starch, methyl
cellulose, agar, betonite, xanthan gum, and the like. Tablets are
formulated, for example, by preparing a powder mixture, granulating or
slugging, adding a lubricant and disintegrant, and pressing into tablets.
A powder mixture is prepared by mixing the compound, suitable comminuted,
with a diluent or base as described above, and optionally, with a binder
such as carboxymethylcellulose, an aliginate, gelating, or polyvinyl
pyrrolidone, a solution retardant such as paraffin, a resorption
accelerator such as a quaternary salt and/or and absorption agent such as
betonite, kaolin, or dicalcium phosphate. The powder mixture can be
granulated by wetting with a binder such as syrup, starch paste, acadia
mucilage, or solutions of cellulosic or polymeric materials and forcing
through a screen. As an alternative to granulating, the powder mixture
can be run through the tablet machine and the result is imperfectly
formed slugs broken into granules. The granules can be lubricated to
prevent sticking to the tablet forming dies by means of the addition of
stearic acid, a stearate salt, talc, or mineral oil. The lubricated
mixture is then compressed into tablets. The compounds of the present
disclosure can also be combined with a free flowing inert carrier and
compressed into tablets directly without going through the granulating or
slugging steps. A clear or opaque protective coating consisting of a
sealing coat of shellac, a coating of sugar or polymeric material, and a
polish coating of wax can be provided. Dyestuffs can be added to these
coatings to distinguish different unit dosages.

[0220] Oral fluids such as solution, syrups, and elixirs can be prepared
in dosage unit form so that a given quantity contains a predetermined
amount of the compound. Syrups can be prepared by dissolving the compound
in a suitably flavored aqueous solution, while elixirs are prepared
through the use of a non-toxic vehicle. Solubilizers and emulsifiers such
as ethoxylated isostearyl alcohols and polyoxyethylene sorbitol ethers,
preservatives, flavor additive such as peppermint oil or natural
sweeteners, or saccharin or other artificial sweeteners, and the like can
also be added.

[0221] Where appropriate, dosage unit formulations for oral administration
can be microencapsulated. The formulation can also be prepared to prolong
or sustain the release as for example by coating or embedding particulate
material in polymers, wax, or the like.

[0222] The compounds of Formula (I), and pharmaceutically acceptable salts
thereof, can also be administered in the form of liposome delivery
systems, such as small unilamellar vesicles, large unilamellar vesicles,
and multilamellar vesicles. Liposomes can be formed from a variety of
phospholipids, such as cholesterol, stearylamine, or phophatidylcholines.

[0223] The compounds of Formula (I) and pharmaceutically acceptable salts
thereof may also be delivered by the use of monoclonal antibodies as
individual carriers to which the compound molecules are coupled. The
compounds may also be coupled with soluble polymers as targetable drug
carriers. Such polymers can include polyvinylpyrrolidone, pyran
copolymer, polyhydroxypropylmethacrylamidephenol,
polyhydroxyethylaspartamidephenol, or polyethyleneoxidepolylysine
substituted with palitoyl residues. Furthermore, the compounds may be
coupled to a class of biodegradable polymers useful in achieving
controlled release of a drug, for example, polylactic acid, polepsilon
caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals,
polydihydropyrans, polycyanoacrylates, and cross-linked or amphipathic
block copolymers of hydrogels.

[0224] Pharmaceutical formulations adapted for transdermal administration
may be presented as discrete patches intended to remain in intimate
contact with the epidermis of the recipient for a prolonged period of
time. For example, the active ingredient may be delivered from the patch
by iontophoresis as generally described in Pharm. Res., 3(6):318 (1986).

[0226] For treatments of the eye or other external tissues, for example
mouth and skin, the formulations are preferably applied as a topical
ointment or cream. When formulated in an ointment, the active ingredient
may be employed with either a paraffinic or a water-miscible ointment
base. Alternatively, the active ingredient may be formulated in a cream
with an oil-in-water cream base or a water-in oil base. Pharmaceutical
formulations adapted for topical administrations to the eye include eye
drops wherein the active ingredient is dissolved or suspended in a
suitable carrier, especially an aqueous solvent.

[0227] Pharmaceutical formulations adapted for topical administration in
the mouth include lozenges, pastilles, and mouth washes.

[0228] Pharmaceutical formulations adapted for rectal administration may
be presented as suppositories or as enemas.

[0229] Pharmaceutical formulations adapted for nasal administration
wherein the carrier is a solid include a course powder having a particle
size for example in the range 20 to 500 microns which is administered in
the manner in which snuff is taken, i.e., by rapid inhalation through the
nasal passage from a container of the powder held close up to the nose.
Suitable formulations wherein the carrier is a liquid, for administration
as a nasal spray or nasal drops, include aqueous or oil solutions of the
active ingredient.

[0230] Pharmaceutical formulations adapted for administration by
inhalation include fine particle dusts or mists, which may be generated
by means of various types of metered, dose pressurized aerosols,
nebulizers, or insufflators.

[0232] Pharmaceutical formulations adapted for parenteral administration
include aqueous and non-aqueous sterile injection solutions which may
contain anti-oxidants, buffers, bacteriostats, and soutes which render
the formulation isotonic with the blood of the intended recipient; and
aqueous and non-aqueous sterile suspensions which may include suspending
agents and thickening agents. The formulations may be presented in
unit-dose or multi-dose containers, for example sealed ampoules and
vials, and may be stored in a freeze-dried (lyophilized) condition
requiring only the addition of the sterile liquid carrier, for example
water for injections, immediately prior to use. Extemporaneous injection
solutions and suspensions may be prepared from sterile powders, granules,
and tablets.

[0233] It should be understood that in addition to the ingredients
particularly mentioned above, the formulations may include other agents
conventional in the art having regard to the type of formulation in
question, for example those suitable for oral administration may include
flavoring agents.

[0234] The term "patient" includes both human and other mammals.

[0235] The term "treating" refers to: (i) preventing a disease, disorder
or condition from occurring in a patient that may be predisposed to the
disease, disorder, and/or condition but has not yet been diagnosed as
having it; (ii) inhibiting the disease, disorder, or condition, i.e.,
arresting its development; and (iii) relieving the disease, disorder, or
condition, i.e., causing regression of the disease, disorder, and/or
condition.

[0236] The compounds of the present disclosure can also be administered
with a cyclosporin, for example, cyclosporin A. Cyclosporin A has been
shown to be active against HCV in clinical trials (Hepatology, 38:1282
(2003); Biochem. Biophys. Res. Commun., 313:42 (2004); J. Gastroenterol.,
38:567 (2003)).

[0237] Table 1 below lists some illustrative examples of compounds that
can be administered with the compounds of this disclosure. The compounds
of the disclosure can be administered with other anti-HCV activity
compounds in combination therapy, either jointly or separately, or by
combining the compounds into a composition.

[0238] The compounds of the present disclosure may also be used as
laboratory reagents. Compounds may be instrumental in providing research
tools for designing of viral replication assays, validation of animal
assay systems and structural biology studies to further enhance knowledge
of the HCV disease mechanisms. Further, the compounds of the present
disclosure are useful in establishing or determining the binding site of
other antiviral compounds, for example, by competitive inhibition.

[0239] The compounds of this disclosure may also be used to treat or
prevent viral contamination of materials and therefore reduce the risk of
viral infection of laboratory or medical personnel or patients who come
in contact with such materials, e.g., blood, tissue, surgical instruments
and garments, laboratory instruments and garments, and blood collection
or transfusion apparatuses and materials.

[0240] This disclosure is intended to encompass compounds having Formula
(I) when prepared by synthetic processes or by metabolic processes
including those occurring in the human or animal body (in vivo) or
processes occurring in vitro.

[0241] The abbreviations used in the present application, including
particularly in the illustrative examples which follow, are well-known to
those skilled in the art. Some of the abbreviations used are as follows:
TFA for trifluoroacetic acid; min or min. or mins for minutes; MeCN or
ACN for acetonitrile; LDA for lithium diisopropylamide; DMSO for
dimethylsulfoxide; h or hr or hrs for hours; Boc or BOC for
tert-butoxycarbonyl; HATU for
O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium
hexafluorophosphate; RT or Rt or rt for retention time or room
temperature (context will dictate); Me for methyl; DMF for
N,N-dimethylformamide; Pd(Ph3P)4 for tetrakistriphenyl
phosphine palladium; MeOH for methanol; MeOD for CD4OD; TEA for
triethylamine; Ph for phenyl; TBDPS for tert-butyldiphenylsilyl;
Et3N or TEA for triethylamine; DMAP for N,N-dimethylaminopyridine;
EtOAc for ethyl acetate; TBAF for tetrabutylammonium fluoride; THF for
tetrahydrofuran; DIEA or DIPEA or iPr2NEt for diisopropylethylamine;
NCS for N-chlorosuccinimide; NBS for N-bromosuccinimide; DCM for
dichloromethane; SEM for 2-(trimethylsilyl)ethoxymethyl; DCE for
1,2-dichloroethane; EDCI for
1-ethyl-3-(3-dimethylaminopropyl)carbodiimide; DBU for
1,8-diazabicyclo[5.4.0]undec-7-ene; Pd(t-Bu3P)2 for palladium
bis(tributylphosphine); HMDS for hexamethyldisilazide; TMSCHN2 for
trimethylsilyldiazomethane; H-D-Ser-OBzl for D-serine benzyl ester;
i-PrOH for isopropanol; LiHMDS for lithium hexamethyldisilazide; DIBAL or
DIBALH for diisobutylaluminum hydride; TBDMS for tert-butyldimethylsilyl;
CBz for carbobenzyloxy; Bn for benzyl; DEAD for diethyl azodicarboxylate;
mCPBA for meta-chloroperoxybenzoic acid; TMSCN for trimethylsilyl
cyanide; dpppe for 1,5-Bis(diphenylphosphino) pentane; TMEDA for
tetramethylethylenediamine; OAc for acetate; DMA for
N,N-dimethylacetamide; and d for days.

[0242] The present disclosure will now be described in connection with
certain embodiments which are not intended to limit its scope. On the
contrary, the present disclosure covers all alternatives, modifications,
and equivalents as can be included within the scope of the claims. Thus,
the following examples, which include specific embodiments, will
illustrate one practice of the present disclosure, it being understood
that the examples are for the purposes of illustration of certain
embodiments and are presented to provide what is believed to be the most
useful and readily understood description of its procedures and
conceptual aspects. Starting materials can be obtained from commercial
sources or prepared by well-established literature methods known to those
of ordinary skill in the art.

EXAMPLES

[0243] Unless noted otherwise, purity assessments were conducted on
Shimadzu LC system, and retention time (Rt) determination and low
resolution mass analysis were conducted on a Shimadzu LC system coupled
with Waters MICROMASS® ZQ MS system. It should be noted that
retention times may vary slightly between machines.

[0403] The optical purity of the two samples of Example D-1b was assessed
according to the chiral HPLC conditions noted below (ee>99% for the
combined crops; ee=96.7% for the sample from flash chromatography):

[0419] Ethynyltrimethylsilane (0.59 mL, 4.25 mmol) was added to a solution
of D-1b (1.5 g, 3.82 mmol), triphenylphosphine (0.20 g, 0.77 mmol),
diethylamine (4.25 mL, 40.70 mmol), copper (I) iodide (40 mg, 0.21 mmol)
and trans-dichloro(bis-triphenylphosphine)palladium (II) (149 mg, 0.21
mmol) in dry DMF (1.4 mL) at rt in a microwave vessel. The vessel was
capped and irradiated for 25 min at 120° C. Two identical 1.5 g
reactions were run in tandem. The reaction mixtures were diluted with
ether and ethyl acetate, combined and shaken with 0.1N HCl. After
standing for 20 min, the suspension was suction-filtered and the pad was
washed with ether and ethyl acetate. The organic phase was then
separated, washed with brine, dried over sodium sulfate, and
concentrated. There was isolated the crude product (4.2 g) as a
brownish-red foam which was taken up in dichloromethane and added
directly to a Thompson 110 g silica gel column. Gradient elution of the
residue with 20% ethyl acetate in dichloromethane to 100% ethyl acetate
furnished D-3d1 (2.8 g, 40% yield) as a yellow solid after evaporation of
the eluant which was taken forward directly. RT=2.37 min (Cond. 3);
LC/MS: Anal. Calcd for C23H32N3O2Si [M+H]+
410.23; found: 410.12.

[0420] D-3d2 to D-3d4 were prepared according to the procedures described
for D-3d1.

[0431] D-3h1 to D-3h6 were prepared from D-3f1, D-3g1, D-3f2, D-3f3, D-3g2
and

[0432] D-3f4, respectively, according to the same procedure used for the
preparation of OL-1e except that methanol (1 mL) was used instead of
dichloromethane. This gave D-3h1 to D-3h6 as hydrochloride salts (or TFA
salts when purified further with preparative HPLC) upon concentration of
the solvent(s) in vacuo.

[0434] Examples D-3 to D-11 were prepared from D-3h1 to D-3h6 and the
appropriate acids according to the same procedure used for the
preparation of Example OL-1. This gave Examples D-3 to D-11 as TFA salts
after HPLC purification.

[0441] Example M2 to M2.1 were prepared as TFA salts from pyrrolidine M1b
and appropriate acids according to the procedure described for the
preparation of Example M1 with a modified purification protocol noted in
the table below.

[0461] Example M3, along with its analogs Examples M4-M7 highlighted in
the table below, were prepared as TFA salts from pyrrolidine M3i (0.4HCl)
by employing the procedure described for the synthesis of Example M1 and
appropriate acids. In the case of Example M7 an equimolar mixture of
(S)-2-(methoxycarbonylamino)-3-methylbutanoic acid and
(S)-2-(methoxycarbonylamino)-2-(tetrahydro-2H-pyran-4-yl)acetic acid was
employed for the coupling step and the resultant statistical mixture of
products was separated by the HPLC technique described for Example M1.
Example M3: LC (Cond. 9 and 10): >95% homogeneity index. LC/MS (Cond.
3): Rt=1.89 min. LC/MS: Anal. Calcd. for
[M+H]+C44H51N8O6: 787.39; found 787.40.

[0463] NCS (0.0195 g, 0.143 mmol) was added to a DMF (2 mL) solution of
the free base form of Example M3 (obtained from the TFA salt via a
standard MCX free-basing protocol; 0.109 g, 0.139 mmol), and stirred at
room temperature for 16 hr and at 50° C. for 25 hr. Most of the
solvent was removed in vacuo, and the residue was dissolved in MeOH and
submitted to a reverse phase HPLC purification (MeOH/water/TFA) to afford
the TFA salt of Example M8 (50 mg) and Example M9 (17.5 mg).

[0474] To a solution of ketone M9.3c (0.4072 g, 1.463 mmol) in THF (7 mL)
was added phenyltrimethylammonium tribromide (1.10 g, 2.93 mmol), and the
reaction mixture was stirred at ˜25° C. for 15 h. The
volatile component was removed in vacuo, and the residue was partitioned
between water (25 mL) and CH2Cl2 (100 mL). The organic layer
was dried over MgSO4, filtered, and concentrated in vacuo to afford
dibromide M9.3d, which was used without further purification. LC/MS:
Anal. Calcd. for [M+H]+C19H1279Br2O2:
434.96; found 434.98.

[0485] Examples M9.5 and M9.6 were prepared as TFA salts starting from
dibromide M9.3b2 according to the procedures described for the
preparation of Examples M9.3 and M9.4 from the corresponding stereoisomer
dibromide M9.3b1.

[0535] To a solution of M12.9b-1 (2.88 g, 11.28 mmol) in Ethanol (20 mL)
was added a solution of LiOH (0.324 g, 13.54 mmol) in water (10.00 mL),
and the mixture was stirred at room temperature for 6 hr. Most of the
volatile component was removed in vacuo, and the residue was partitioned
between water (20 mL) and ether (20 mL). The aqueous layer was chilled in
an ice-water bath, acidified with a 1N HCl to a pH region of 2, and
extracted with EtOAc (30 mL, 4×). The combined organic phase was
dried with Na2SO4, evaporated in vacuo to give acid M12.9c as a
sticky solid (2.55 g). 1H NMR (CDCl3, 400 MHz): 4.64 (m, 1H),
3.25 (appt s, 1H), 2.70-2.40 (m, 1H), 2.14 (m, 1H), 1.54-1.44 (m, 9H),
1.27 (s, 3H), 1.10-0.80 (m, 1H), 0.67 (m, 1H).

Example M12.9, Step d

##STR00220##

[0537] To a solution of acid M12.9c (2.05 g, 8.50 mmol) in Acetonitrile
(50 mL) was added 2-bromo-1-(4-bromophenyl)ethanone (2.361 g, 8.50 mmol)
followed by DIEA (1.484 mL, 8.50 mmol), and the reaction mixture was
stirred at room temperature for 16 hr. Most of the volatile component was
removed in vacuo, and the residue was partitioned between EtOAc (50 mL)
and water (50 mL). The organic layer was washed with sat. NaHCO3 (30
mL) and sat. NaCl (20 mL), dried with Na2SO4, and evaporated in
vacuo to afford ketoester M12.9d as white foam (3.5 g). LC/MS: Anal.
Calcd. for [M+Na]+C20H2481BrNNaO5: 462.07;
found: 461.91.

[0541] To a pressure tube containing a solution of bromide M12.9e (0.354
g, 1.007 mmol) and 1,2-bis(trimethylstannyl)ethyne (0.354 g, 1.007 mmol)
in DMF (15 mL) was added Pd (Ph3P)4 (0.070 g, 0.060 mmol), and
the reaction mixture was degassed for 10 min and the reaction vessel was
capped and heated at 90° C. for 14 hr. Most of the volatile
component was removed in vacuo, and the residue was partitioned between
DCM (60 mL) and water (40 mL). The organic layer was dried with
Na2SO4, and evaporated in vacuo. The resulting crude material
was purified with flash chromatograph (40-100% EtOAc/hexanes) to afford
alkyne M12.9f as a red solid (0.3 g). LC/MS: Anal. Calcd. for
[M+H]+C42H49N6O4: 701.38; found: 701.43.

Example M12.9, Step g

##STR00223##

[0543] 4N HCl in dioxane (3.90 mL, 128 mmol) was added to carbamate M12.9f
(0.3 g, 0.428 mmol), and the mixture was stirred at room temperature for
5 hr. The volatile component was removed in vacuo and the residue was
dried under high vacuum overnight to afford the HCl salt of pyrrolidine
M12.9 g as a yellow solid (0.27 g). LC/MS: Anal. Calcd. for
[M+H]+C32H33N6: 501.28; found: 501.22.

[0555] Example M14 (TFA salt) was prepared from pyrrolidine M13d and
appropriate acid by employing the procedure described for the synthesis
of Example M1. In the case of Example M15 an equimolar mixture of
(S)-2-(methoxycarbonylamino)-3-methylbutanoic acid and
(S)-2-(methoxycarbonylamino)-2-(tetrahydro-2H-pyran-4-yl)acetic acid was
employed for the coupling step and the resultant statistical mixture of
products was separated by the HPLC technique described for Example M1.

[0566] In a sealed tube, a mixture of ketoamide N1d (0.12 g, 0.158 mmol)
and ammonium acetate (0.122 g, 1.582 mmol) in xylene (2 mL) was heated at
140° C. for 4 hrs. The reaction mixture was partitioned between
EtOAc and water, the organic layer was washed with sat. NaHCO3 and
sat. NaCl, dried over anhydrous Na2SO4, filtered and
concentrated. The residue was dissolved in a small amount of methylene
chloride and charged to a 40 g silica gel cartridge which was eluted with
a 20 min gradient of 0-100% EtOAc in hexane. Imidazole N1e (0.054 g) was
collected as a yellow solid. LC/MS (Cond. 10d): Rt=3.3 min. LC/MS:
Anal. Calcd. For [M+H]+ C38H41F4N6O4:
721.3; found: 721.10.

[0572] Example N5-N7.3 (TFA salt) were prepared starting from aminoketone
N1c and appropriate starting materials, obtained from commercial sources,
by employing the procedures described for the synthesis of Example N1.

[0580] Compound N9 (TFA salt) was prepared from pyrrolidine N8a and
(S)-2-(methoxycarbonylamino)-2-(tetrahydro-2H-pyran-4-yl)acetic acid
according to the procedure described for the preparation of Example N1
with the exception that ACN/water/TFA solvent system was employed for the
purification step. LC (Cond. 10e): >97% homogeneity index. LC/MS
(Cond. 10g): Rt=1.82 min. LC/MS: Anal. Calcd. for
[M+H]+C48H55N8O8: 871.41; found 871.6.

Example N10

##STR00253##

[0581] Example N10, Step a

##STR00254##

[0583] A solution of (S)-5-oxopyrrolidine-2-carboxylic acid (20 g, 154
mmol) in MeOH (140 mL) was cooled to 0° C. Then thionyl chloride
(11.2 mL, 154 mmol) was added drop wise and stirred for 30 minutes. The
reaction mixture was brought to RT and stirred for 3 h. Then the reaction
mixture was concentrated under reduced pressure, the resulting residue
was dissolved in EtOAc (200 mL) and the organic layer was washed with
NaHCO3 solution. The aqueous phase was extracted with EtOAc
(2×100 mL), and the combined organic layer was washed with brine,
dried over Na2SO4 and concentrated in vacuo to obtain crude
ester N10a (8.0 g), which was used as such in the next step. 1H NMR
(DMSO-d6, δ=2.50 ppm, 400 MHz): δ 7.99 (s, 1H),
4.21-4.17 (m, 1H), 3.67 (s, 3H), 2.39-2.29 (m, 1H), 2.17-2.09 (m, 2H),
2.02-1.95 (m, 1H).

[0596] Example N11 (TFA salt) was prepared from pyrrolidine if (4HCl salt)
and (S)-2-(methoxycarbonylamino)-2-(tetrahydro-2H-pyran-4-yl)acetic acid
according to the procedure described for Example N10 with the exception
that ACN/water/TFA was used for the HPLC purification step. LC (Condition
10e and 10e.1): >98% homogeneity index. LC/MS (Condition 10e.5):
Rt=1.44 min. LC/MS: Anal. Calcd. for
[M+H]+C48H59N8O8: 875.44; found 875.4.

[0617] A solution of ester N14a (1.4 g, 5.16 mmol) in THF (25 mL) was
cooled to -78° C. and Superhydride (1M in THF, 6.19 mL, 6.19 mmol)
was added to it drop wise under the N2. The reaction mixture was
stirred for 30 minutes, quenched with saturated NaHCO3 solution and
allowed to warm up to 0° C. Then 30% aqueous H2O2 (3 mL)
was added and stirred for 20 minutes at 0° C. The reaction mixture
was concentrated under reduced pressure and the residue was dissolved in
DCM (100 mL) and washed with water. The aqueous phase was extracted with
DCM (2×50 mL), and the combined organic extract was washed with
brine, dried over Na2SO4 and concentrated in vacuo to afford
crude N14b (1.25 g) which was used as such in the next step.

[0676] TFA (3 mL, 38.9 mmol) was added to a stirred solution of carbamate
J1c (238 mg, 0.367 mmol) in DCE (7 mL) and the reaction was stirred at rt
for 1 h. The reaction mixture was concentrated under vacuum to yield a
TFA salt of pyrrolidine J1d (260 mg) as a yellow solid.

[0689] TFA (3 mL, 38.9 mmol) was added to a stirred solution of carbamate
J3b (316 mg, 0.470 mmol) in DCE (7 mL) and the reaction was stirred at rt
for 1 h. The reaction mixture was concentrated under vacuum to yield a
TFA salt of pyrrolidine J3c (360 mg) as a yellow solid. LC-MS retention
time 2.711 min; m/z 473.24 (MH+). LC data was recorded on a Shimadzu
LC-10AS liquid chromatograph equipped with a Phenomenex-Luna 10u C18
3.0×50 mm column using a SPD-10AV UV-Vis detector at a detector
wave length of 220 nM. The elution conditions employed a flow rate of 4
mL/min, a gradient of 100% solvent A/0% solvent B to 0% solvent A/100%
solvent B, a gradient time of 3 min, a hold time of 1 min, and an
analysis time of 4 min where solvent A was 5% MeOH/95% H2O/10 mM
ammonium acetate and solvent B was 5% H2O/95% MeOH/10 mM ammonium
acetate. MS data was determined using a Micromass Platform for LC in
electrospray mode.

[0697] TFA (250 μL, 3.24 mmol) was added to a solution of carbamate J5a
in DCE (1 mL) and the reaction was stirred at rt for 1.5 hr. The reaction
was concentrated under a stream of nitrogen to provide a TFA salt of
pyrrolidine J5b (45 mg). LC-MS retention time 2.928 min; m/z 525.32
(MH+). LC data was recorded on a Shimadzu LC-10AS liquid chromatograph
equipped with a Phenomenex-Luna 10u C18 3.0×50 mm column using a
SPD-10AV UV-Vis detector at a detector wave length of 220 nM. The elution
conditions employed a flow rate of 4 mL/min, a gradient of 100% solvent
A/0% solvent B to 0% solvent A/100% solvent B, a gradient time of 4 min,
a hold time of 1 min, and an analysis time of 5 min where solvent A was
5% MeOH/95% H2O/10 mM ammonium acetate and solvent B was 5%
H2O/95% MeOH/10 mM ammonium acetate. MS data was determined using a
Micromass Platform for LC in electrospray mode.

[0699] Compound analysis conditions: Purity assessment and low resolution
mass analysis were conducted on a Shimadzu LC system coupled with Waters
Micromass ZQ MS system. It should be noted that retention times may vary
slightly between machines. Additional LC conditions applicable to the
current section, unless noted otherwise.

[0726] NaBH3CN (6.22 g, 94 mmol) was added in portions over a few
minutes to a cooled (ice/water) mixture of (R)-2-Phenylglycine (6.02 g,
39.8 mmol) and methanol (100 mL), and stirred for 5 minutes. Acetaldehyde
(10 mL) was added dropwise over 10 minutes and stirring was continued at
the same cooled temperature for 45 minutes and at ambient temperature for
˜6.5 hours. The reaction mixture was cooled back with ice-water
bath, treated with water (3 mL) and then quenched with a dropwise
addition of concentrated HCl over ˜45 minutes until the pH of the
mixture was ˜1.5-2.0. The cooling bath was removed and the stirring
was continued while adding concentrated HCl in order to maintain the pH
of the mixture around 1.5-2.0. The reaction mixture was stirred
overnight, filtered to remove the white suspension, and the filtrate was
concentrated in vacuo. The crude material was recrystallized from ethanol
to afford the HCl salt of Cap-2 as a shining white solid in two crops
(crop-1: 4.16 g; crop-2: 2.19 g). 1H NMR (DMSO-d6, δ=2.5
ppm, 400 MHz): 10.44 (1.00, br s, 1H), 7.66 (m, 2H), 7.51 (m, 3H), 5.30
(s, 1H), 3.15 (br m, 2H), 2.98 (br m, 2H), 1.20 (app br s, 6H). Crop-1:
[α]25-102.21° (c=0.357, H2O); crop-2:
[α]25-99.7° (c=0.357, H2O). LC (Cond. I): RT=0.43
min; LC/MS: Anal. Calcd. for [M+H]+ C12H18NO2:
208.13; found 208.26.

[0740] A methanol (10 mL) solution of either enantiomer of benzyl
2-(4-methylpiperazin-1-yl)-2-phenylacetate (1.0 g, 3.1 mmol) was added to
a suspension of 10% Pd/C (120 mg) in methanol (5.0 mL). The reaction
mixture was exposed to a balloon of hydrogen, under a careful monitoring,
for <50 minutes. Immediately after the completion of the reaction, the
catalyst was filtered through diatomaceous earth (Celite®) and the
filtrate was concentrated in vacuo to provide Cap-7, contaminated with
phenylacetic acid as a tan foam (867.6 mg; mass is above the theoretical
yield). The product was used for the next step without further
purification. 1H NMR (DMSO-d6, δ=2.5, 500 MHz) δ
7.44-7.37 (m, 2H), 7.37-7.24 (m, 3H), 3.92 (s, 1H), 2.63-2.48 (app. br s,
2H), 2.48-2.32 (m, 6H), 2.19 (s, 3H); RT=0.31 (Cond. II); >90%
homogeneity index; LC/MS: Anal. Calcd. for
[M+H]+C13H19N2O2: 235.14; found 235.15; HRMS:
Anal. Calcd. for [M+H]+C13H19N2O2: 235.1447;
found 235.1440.

[0741] The synthesis of Cap-8 and Cap-9 was conducted according to the
synthesis of Cap-7 by using appropriate amines for the SN2
displacement step (i.e., 4-hydroxypiperidine for Cap-8 and
(S)-3-fluoropyrrolidine for Cap-9) and modified conditions for the
separation of the respective stereoisomeric intermedites, as described
below.

[0756] Step 2: To a stirred solution of the intermediate ester (1.12 g,
2.88 mmol) in dichloromethane (10 mL) was added TFA (3 mL). The reaction
mixture was stirred at ambient temperature for 4 hours and then it was
concentrated to dryness to give a light yellow oil. The oil was purified
using reverse-phase preparative HPLC (Primesphere C-18, 30×100 mm;
CH3CN--H2O-0.1% TFA). The appropriate fractions were combined
and concentrated to dryness in vacuo. The residue was then dissolved in a
minimum amount of methanol and applied to applied to MCX LP extraction
cartridges (2×6 g). The cartridges were rinsed with methanol (40
mL) and then the desired compound was eluted using 2M ammonia in methanol
(50 mL). Product-containing fractions were combined and concentrated and
the residue was taken up in water. Lyophilization of this solution
provided the title compound (0.492 g, 78%) as a light yellow solid.
1H NMR (DMSO-d6) δ 7.50 (s, 5H), 5.13 (s, 1H), 3.09 (br
s, 2H), 2.92-2.89 (m, 2H), 1.74 (m, 4H), 1.48 (br s, 2H). LC/MS: Anal.
Calcd. for C13H17NO2: 219; found: 220 (M+H)+.

[0785] Step 3; (R,S)-2-(4-Pyridyl)-2-(N,N-dimethylamino)acetic acid: To a
solution of (R,S)-ethyl 2-(4-pyridyl)-2-(N,N-dimethylamino)acetate (0.200
g, 0.960 mmol) in a mixture of THF-methanol-H2O (1:1:1, 6 mL) was
added powdered LiOH (0.120 g, 4.99 mmol) at room temperature. The
solution was stirred for 3 hours and then it was acidified to pH 6 using
1N HCl. The aqueous phase was washed with ethyl acetate and then it was
lyophilized to give the dihydrochloride of the title compound as a yellow
solid (containing LiCl). The product was used as such in subsequent
steps. 1H NMR (400 MHz, DMSO-d6) δ 8.49 (d, J=5.7 Hz,
2H), 7.34 (d, J=5.7 Hz, 2H), 3.56 (s, 1H), 2.21 (s, 6H).

[0786] The following examples were prepared in similar fashion using the
method described above;

[0789] Step 2; (R,S) 2-(Quinolin-3-yl)-2-(N,N-dimethylamino)acetic acid: A
mixture of (R,S)-ethyl 2-(quinolin-3-yl)-2-(N,N-dimethylamino)acetate
(0.122 g, 0.472 mmol) and 6M HCl (3 mL) was heated at 100° C. for
12 hours. The solvent was removed in vacuo to provide the dihydrochloride
of the title compound (0.169 g, >100%) as a light yellow foam. The
unpurified material was used in subsequent steps without further
purification. LCMS: Anal. Calcd. for C13H14N2O2: 230;
found: 231 (M+H)+.

[0839] NaH (0.0727 g, 1.82 mmol, 60%) was added to a cooled (ice-water)
THF (3.0 mL) solution of the TFA salt (R)-benzyl
2-(diethylamino)-3-hydroxypropanoate (0.3019 g, 0.8264 mmol) prepared
above, and the mixture was stirred for 15 min. Methyl iodide (56 μL,
0.90 mmol) was added and stirring was continued for 18 hr while allowing
the bath to thaw to ambient condition. The reaction was quenched with
water and loaded onto a MeOH pre-conditioned MCX (6 g) cartridge, and
washed with methanol followed by compound elution with 2N
NH3/Methanol. Removal of the volatile component in vacuo afforded
Cap-75, contaminated with (R)-2-(diethylamino)-3-hydroxypropanoic acid,
as a yellow semi-solid (100 mg). The product was used as is without
further purification.

Cap-76

##STR00398##

[0841] NaCNBH3 (1.60 g, 24.2 mmol) was added in batches to a chilled
(˜15° C.) water/MeOH (12 mL each) solution of
(S)-4-amino-2-(tert-butoxycarbonylamino) butanoic acid (2.17 g, 9.94
mmol). A few minutes later acetaldehyde (2.7 mL, 48.1 mmol) was added
drop-wise over 2 min, the cooling bath was removed, and the reaction
mixture was stirred at ambient condition for 3.5 hr. An additional
acetaldehyde (2.7 mL, 48.1 mmol) was added and the reaction was stirred
for 20.5 hr. Most of the MeOH component was removed in vacuo, and the
remaining mixture was treated with concentrated HCl until its pH reached
˜1.0 and then heated for 2 hr at 40° C. The volatile
component was removed in vacuo, and the residue was treated with 4 M
HCl/dioxane (20 mL) and stirred at ambient condition for 7.5 hr. The
volatile component was removed in vacuo and the residue was purified with
Dowex® 50WX8-100 ion-exchange resin (column was washed with water and
the compound was eluted with dilute NH4OH, prepared from 18 ml of
NH4OH and 282 ml of water) to afford intermediate
(S)-2-amino-4-(diethylamino)butanoic acid as an off-white solid (1.73 g).

[0842] Methyl chloroformate (0.36 mL, 4.65 mmol) was added drop-wise over
11 min to a cooled (ice-water) mixture of Na2CO3 (0.243 g, 2.29
mmol), NaOH (4.6 mL of 1M/H2O, 4.6 mmol) and the above product
(802.4 mg). The reaction mixture was stirred for 55 min, and then the
cooling bath was removed and stirring was continued for an additional
5.25 hr. The reaction mixture was diluted with equal volume of water and
washed with CH2Cl2 (30 mL, 2×), and the aqueous phase was
cooled with ice-water bath and acidified with concentrated HCl to a pH
region of 2. The volatile component was then removed in vacuo and the
crude material was free-based with MCX resin (6.0 g; column was washed
with water, and sample was eluted with 2.0 M NH3/MeOH) to afford
impure Cap-76 as an off-white solid (704 mg). 1H NMR (MeOH-d4,
δ=3.29 ppm, 400 MHz): δ 3.99 (dd, J=7.5, 4.7, 1H), 3.62 (s,
3H), 3.25-3.06 (m, 6H), 2.18-2.09 (m, 1H), 2.04-1.96 (m, 1H), 1.28 (t,
J=7.3, 6H). LC/MS: Anal. Calcd. for
[M+H]+C10H21N2O4: 233.15; found 233.24.

Cap-77a and -77b

##STR00399##

[0844] The synthesis of Cap-77 was conducted according to the procedure
described for Cap-7 by using 7-azabicyclo[2.2.1]heptane for the SN2
displacement step, and by effecting the enantiomeric separation of the
intermediate benzyl 2-(7-azabicyclo[2.2.1]heptan-7-yl)-2-phenylacetate
using the following condition: the intermediate (303.7 mg) was dissolved
in ethanol, and the resulting solution was injected on a chiral HPLC
column (Chiracel AD-H column, 30×250 mm, 5 um) eluting with 90%
CO2-10% EtOH at 70 mL/min, and a temperature of 35° C. to
provide 124.5 mg of enantiomer-1 and 133.8 mg of enantiomer-2. These
benzyl esters were hydrogenolysed according to the preparation of Cap-7
to provide Cap-77: 1H NMR (DMSO-d6, δ=2.5 ppm, 400 MHz):
δ 7.55 (m, 2H), 7.38-7.30 (m, 3H), 4.16 (s, 1H), 3.54 (app br s,
2H), 2.08-1.88 (m, 4H), 1.57-1.46 (m, 4H). LC (Cond. 1): RT=0.67 min;
LC/MS: Anal. Calcd. for [M+H]+C14H18NO2: 232.13;
found 232.18. HRMS: Anal. Calcd. for [M+H]+C14H18NO2:
232.1338; found 232.1340.

Cap-78

##STR00400##

[0846] NaCNBH3 (0.5828 g, 9.27 mmol) was added to a mixture of the
HCl salt of (R)-2-(ethylamino)-2-phenylacetic acid (an intermediate in
the synthesis of Cap-3; 0.9923 mg, 4.60 mmol) and
(1-ethoxycyclopropoxy)trimethylsilane (1.640 g, 9.40 mmol) in MeOH (10
mL), and the semi-heterogeneous mixture was heated at 50° C. with
an oil bath for 20 hr. More (1-ethoxycyclopropoxy)trimethylsilane (150
mg, 0.86 mmol) and NaCNBH3 (52 mg, 0.827 mmol) were added and the
reaction mixture was heated for an additional 3.5 hr. It was then allowed
to cool to ambient temperature and acidified to a˜pH region of 2
with concentrated HCl, and the mixture was filtered and the filtrate was
rotervaped. The resulting crude material was taken up in i-PrOH (6 mL)
and heated to effect dissolution, and the non-dissolved part was filtered
off and the filtrate concentrated in vacuo. About 1/3 of the resultant
crude material was purified with a reverse phase HPLC(H2O/MeOH/TFA)
to afford the TFA salt of Cap-78 as a colorless viscous oil (353 mg).
1H NMR (DMSO-d6, δ=2.5 ppm, 400 MHz; after D2O
exchange): δ 7.56-7.49 (m, 5H), 5.35 (S, 1H), 3.35 (m, 1H), 3.06
(app br s, 1H), 2.66 (m, 1H), 1.26 (t, J=7.3, 3H), 0.92 (m, 1H),
0.83-0.44 (m, 3H). LC (Cond. 1): RT=0.64 min; LC/MS: Anal. Calcd. for
[M+H]+ C13H18NO2: 220.13; found 220.21. HRMS: Anal.
Calcd. for [M+H]+C13H18NO2: 220.1338; found 220.1343.

Cap-79

##STR00401##

[0848] Ozone was bubbled through a cooled (-78° C.)
CH2Cl2 (5.0 mL) solution Cap-55 (369 mg, 2.13 mmol) for about
50 min until the reaction mixture attained a tint of blue color.
Me2S (10 pipet drops) was added, and the reaction mixture was
stirred for 35 min. The -78° C. bath was replaced with
a-10° C. bath and stirring continued for an additional 30 min, and
then the volatile component was removed in vacuo to afford a colorless
viscous oil.

[0849] NaBH3CN (149 mg, 2.25 mmol) was added to a MeOH (5.0 mL)
solution of the above crude material and morpholine (500 μL, 5.72
mmol) and the mixture was stirred at ambient condition for 4 hr. It was
cooled to ice-water temperature and treated with concentrated HCl to
bring its pH to ˜2.0, and then stirred for 2.5 hr. The volatile
component was removed in vacuo, and the residue was purified with a
combination of MCX resin (MeOH wash; 2.0 N NH3/MeOH elution) and a
reverse phase HPLC(H2O/MeOH/TFA) to afford Cap-79 containing unknown
amount of morpholine.

[0850] In order to consume the morpholine contaminant, the above material
was dissolved in CH2Cl2 (1.5 mL) and treated with Et3N
(0.27 mL, 1.94 mmol) followed by acetic anhydride (0.10 mL, 1.06 mmol)
and stirred at ambient condition for 18 hr. THF (1.0 mL) and H2O
(0.5 mL) were added and stirring continued for 1.5 hr. The volatile
component was removed in vacuo, and the resultant residue was passed
through MCX resin (MeOH wash; 2.0 N NH3/MeOH elution) to afford
impure Cap-79 as a brown viscous oil, which was used for the next step
without further purification.

[0861] Prepared according to the protocol described by Falb et al.
Synthetic Communications 1993, 23, 2839.

Cap-82 to Cap-85

[0862] Cap-82 to Cap-85 were synthesized from appropriate starting
materials according to the procedure described for Cap-51 or Cap-13. The
samples exhibited similar spectral profiles as that of their enantiomers
(i.e., Cap-4, Cap-13, Cap-51 and Cap-52, respectively).

[0870] A mixture of L-valine (1.0 g, 8.54 mmol), 5-bromopyrimidine (4.03
g, 17.0 mmol), K2CO3 (2.40 g, 17.4 mmol) and Cut (179 mg, 0.94
mmol) in DMSO (10 mL) was heated at 100° C. for 12 h. The reaction
mixture was cooled to RT, poured into H2O (ca. 150 mL) and washed
with EtOAc (×2). The organic layers were extracted with a small
amount of H2O and the combined aq phases were acidified to ca. pH 2
with 6N HCl. The volume was reduced to about one-third and 20 g of cation
exchange resin (Strata) was added. The slurry was allowed to stand for 20
min and loaded onto a pad of cation exchange resin (Strata) (ca. 25 g).
The pad was washed with H2O (200 mL), MeOH (200 mL), and then
NH3 (3M in MeOH, 2×200 mL). The appropriate fractions was
concentrated in vacuo and the residue (ca. 1.1 g) was dissolved in
H2O, frozen and lyophyllized. The title compound was obtained as a
foam (1.02 g, 62%). 1HNMR (400 MHz, CD3OD) showed the mixture
to contain valine and the purity could not be estimated. The material was
used as is in subsequent reactions. LCMS: Anal. Calcd. for
C9H13N3O2: 195; found: 196 (M+H)+.

Cap-90

##STR00409##

[0872] Cap-90 was prepared according to the method described for the
preparation of Cap-1. The crude material was used as is in subsequent
steps. LCMS: Anal. Calcd. for C11H15NO2: 193; found: 192
(M-H)-.

[0873] The following caps were prepared according to the method used for
preparation of cap 51 unless noted otherwise:

[0874] For the preparation of Cap-117 to Cap-123 the Boc amino acids were
obtained from commercially sources and were deprotected by treatment with
25% TFA in CH2Cl2. After complete reaction as judged by LCMS
the solvents were removed in vacuo and the corresponding TFA salt of the
amino acid was carbamoylated with methyl chloroformate according to the
procedure described for Cap-51.

[0876] The hydrochloride salt of L-threonine tert-butyl ester was
carbamoylated according to the procedure for Cap-51. The crude reaction
mixture was acidified with 1N HCl to pH˜1 and the mixture was
extracted with EtOAc (2×50 mL). The combined organic phases were
concentrated in vacuo to give a colorless oil which solidified on
standing. The aqueous layer was concentrated in vacuo and the resulting
mixture of product and inorganic salts was triturated with
EtOAc--CH2Cl2-MeOH (1:1:0.1) and then the organic phase
concentrated in vacuo to give a colorless oil which was shown by LCMS to
be the desired product. Both crops were combined to give 0.52 g of a
solid. 1HNMR (400 MHz, CD3OD) δ 4.60 (m, 1H), 4.04 (d,
J=5.0 Hz, 1H), 1.49 (d, J=6.3 Hz, 3H). LCMS: Anal. Calcd. for
C5H7NO4: 145; found: 146 (M+H)+.

Cap-125

##STR00445##

[0878] To a suspension of Pd(OH)2, (20%, 100 mg), aqueous
formaldehyde (37% wt, 4 ml), acetic acid, (0.5 mL) in methanol (15 mL)
was added (S)-4-amino-2-(tert-butoxycarbonylamino)butanoic acid (1 g,
4.48 mmol). The reaction was purged several times with hydrogen and was
stirred overnight with an hydrogen balloon room temp. The reaction
mixture was filtered through a pad of diatomaceous earth (Celite®),
and the volatile component was removed in vacuo. The resulting crude
material was used as is for the next step. LC/MS: Anal. Calcd. for
C11H22N2O4: 246; found: 247 (M+H)+.

Cap-126

##STR00446##

[0880] This procedure is a modification of that used to prepare Cap-51. To
a suspension of 3-methyl-L-histidine (0.80 g, 4.70 mmol) in THF (10 mL)
and H2O (10 mL) at 0° C. was added NaHCO3 (0.88 g, 10.5
mmol). The resulting mixture was treated with C1CO2Me (0.40 mL, 5.20
mmol) and the mixture allowed to stir at 0° C. After stirring for
ca. 2h LCMS showed no starting material remaining. The reaction was
acidified to pH 2 with 6 N HCl.

[0881] The solvents were removed in vacuo and the residue was suspended in
20 mL of 20% MeOH in CH2Cl2. The mixture was filtered and
concentrated to give a light yellow foam (1.21 g). LCMS and 1H NMR
showed the material to be a 9:1 mixture of the methyl ester and the
desired product. This material was taken up in THF (10 mL) and H2O
(10 mL), cooled to 0° C. and LiOH (249.1 mg, 10.4 mmol) was added.
After stirring ca. 1 h LCMS showed no ester remaining. Therefore the
mixture was acidified with 6N HCl and the solvents removed in vacuo. LCMS
and 1H NMR confirm the absence of the ester. The title compound was
obtained as its HCl salt contaminated with inorganic salts (1.91 g,
>100%). The compound was used as is in subsequent steps without
further purification. 1HNMR (400 MHz, CD3OD) δ 8.84, (s,
1H), 7.35 (s, 1H), 4.52 (dd, J=5.0, 9.1 Hz, 1H), 3.89 (s, 3H), 3.62 (s,
3H), 3.35 (dd, J=4.5, 15.6 Hz, 1H, partially obscured by solvent), 3.12
(dd, J=9.0, 15.6 Hz, 1H).LCMS: Anal. Calcd. for
C9H13N3O4: 227.09; found: 228.09 (M+H)+.

[0906] To a cooled (-50° C.) suspension of 1-benzyl-1H-imidazole
(1.58 g, 10.0 mmol) in anhydrous diethyl ether (50 mL) under nitrogen was
added n-butyl lithium (2.5 M in hexanes, 4.0 mL, 10.0 mmol) dropwise.
After being stirred for 20 min at -50° C., dry carbon dioxide
(passed through Drierite) was bubbled into the reaction mixture for 10
min before it was allowed to warm up to 25° C. The heavy
precipitate which formed on addition of carbon dioxide to the reaction
mixture was filtered to yield a hygroscopic, white solid which was taken
up in water (7 mL), acidified to pH=3, cooled, and induced to crystallize
with scratching. Filtration of this precipitate gave a white solid which
was suspended in methanol, treated with 1N HCl/diethyl ether (4 mL) and
concentrated in vacuo. Lyophilization of the residue from water (5 mL)
afforded the HCl salt of Cap-136 as a white solid (817 mg, 40%). 1H
NMR (300 MHz, DMSO-d6) δ 7.94 (d, J=1.5 Hz, 1H), 7.71 (d,
J=1.5 Hz, 1H), 7.50-7.31 (m, 5H), 5.77 (s, 2H); Rt=0.51 min
(Cond.-MS-W5); 95% homogenity index; LRMS: Anal. Calc. for
[M+H]+C11H12N2O2: 203.08; found: 203.11.

[0927] To a vigorously-stirred mixture of
1,3-dichloro-5-ethoxyisoquinoline (482 mg, 2.00 mmol; prepared according
to the procedure in WO 2005/051410), palladium (II) acetate (9 mg, 0.04
mmol), sodium carbonate (223 mg, 2.10 mmol) and
1,5-bis(diphenylphosphino)pentane (35 mg, 0.08 mmol) in dry
dimethylacetamide (2 mL) at 25° C. under nitrogen was added
N,N,N',N'-tetramethylethylenediamine (60 mL, 0.40 mmol). After 10 min,
the mixture was heated to 150° C., and then a stock solution of
acetone cyanohydrin (prepared from 457 μL of acetone cyanohydrin in
4.34 mL DMA) was added in 1 mL portions over 18 h using a syringe pump.
The mixture was then partitioned between ethyl acetate and water and the
organic layer was separated, washed with brine, dried over
Na2SO4, filtered and concentrated. The residue was purified on
silica gel (gradient elution with 10% ethyl acetate in hexanes to 40%
ethyl acetate in hexanes) to afford Cap-140, step a (160 mg, 34%) as a
yellow solid. Rt=2.46 min (Cond.-MS-W2); 90% homogenity index; LCMS:
Anal. Calc. for [M+H]+C12H9ClN2O: 233.05; found:
233.08.

Cap-140

[0928] Cap-140 was prepared by the acid hydrolysis of Cap-140, step a with
12N HCl as described in the procedure for the preparation of Cap 141,
described below. Rt=2.24 min (Cond.-MS-W2); 90% homogenity index;
LCMS: Anal. Calc. for [M+H]+C12H11ClNO3: 252.04;
found: 252.02.

[0941] To a stirred solution of 3-amino-1-bromoisoquinoline (444 mg, 2.00
mmol) in anhydrous dimethylformamide (10 mL) was added sodium hydride
(60%, unwashed, 96 mg, 2.4 mmol) in one portion. The mixture was stirred
at 25° C. for 5 min before 2-bromoethyl ether (90%, 250 μL,
2.00 mmol) was added. This mixture was stirred further at 25° C.
for 5 h and at 75° C. for 72 h before it was cooled to 25°
C., quenched with saturated ammonium chloride solution and diluted with
ethyl acetate. The organic layer was separated, washed with water and
brine, dried over Na2SO4, filtered and concentrated.
Purification of the residue on silica gel (gradient elution with 0% to
70% ethyl acetate in hexanes) afforded Cap-143, step a (180 mg, 31%) as a
yellow solid. Rt=1.75 min (Cond.-MS-W1); 90% homogenity index; LCMS:
Anal. Calc. for [M+1-1]+C13H14BrN2O: 293.03; found:
293.04.

Cap-143

[0942] To a cold (-60° C.) solution of Cap-143, step a (154 mg,
0.527 mmol) in anhydrous tetrahydrofuran (5 mL) was added a solution of
n-butyllithium in hexanes (2.5 M, 0.25 mL, 0.633 mmol). After 10 min, dry
carbon dioxide was bubbled into the reaction mixture for 10 min before it
was quenched with 1N HCl and allowed to warm to 25° C. The mixture
was then extracted with dichloromethane (3×30 mL) and the combined
organic extracts were concentrated in vacuo. Purification of the residue
by reverse phase HPLC (MeOH/water/TFA) afforded Cap-143 (16 mg, 12%).
Rt=1.10 min (Cond.-MS-W1); 90% homogenity index; LCMS: Anal. Calc.
for [M+H]+C14H15N2O3: 259.11; found: 259.08.

Cap-144

##STR00482##

[0943] Cap-144, Step a

##STR00483##

[0945] 1,3-Dichloroisoquinoline (2.75 g, 13.89 mmol) was added in small
portions to a cold (0° C.) solution of fuming nitric acid (10 mL)
and concentrated sulfuric acid (10 mL). The mixture was stirred at
0° C. for 0.5 h before it was gradually warmed to 25° C.
where it stirred for 16 h. The mixture was then poured into a beaker
containing chopped ice and water and the resulting suspension was stirred
for 1 h at 0° C. before it was filtered to afford Cap-144, step a
(2.73 g, 81%) as a yellow solid which was used directly. Rt=2.01 min
(Cond.-D1); 95% homogenity index; LCMS: Anal. Calc. for
[M+H]+C9H5Cl2N2O2: 242.97; found: 242.92.

[0950] Cap-144 was prepared according to the procedure described for
Cap-141. Rt=2.36 min (Cond.-D1); 90%; LCMS: Anal. Calc. for
[M+H]+C12H12ClN2O2: 238.01; found: 238.09.

Caps-145 to -162 Caps-145 to 162 were prepared from the appropriate
1-chloroisoquinolines according to the procedure described for the
preparation of Cap-138 (Method A) or Cap-139 (Method B) unless noted
otherwise as outlined below.

[0978] The following diazotization step was adapted from Barton, A.;
Breukelman, S. P.; Kaye, P. T.; Meakins, G. D.; Morgan, D. J. J. C. S.
Perkin Trans I 1982, 159-164: A solution of NaNO2 (150 mg, 2.17
mmol) in water (1.0 mL) was added dropwise to a stirred, cold (0°
C.) solution of methyl 2-amino-5-ethyl-1,3-thiazole-4-carboxylate (186
mg, 1.0 mmol) in 50% H3PO2 (3.2 mL). The mixture was stirred at
0° C. for 1 h and allowed to warm up to room temperature where it
stirred further for 2 h. After recooling to 0° C., the mixture was
treated slowly with a solution of NaOH (85 mg) in water (10 mL). The
mixture was then diluted with saturated NaHCO3 solution and
extracted twice with ether. The organic layers were combined, dried over
MgSO4 and concentrated to give methyl 5-ethylthiazole-4-carboxylate
(i.e. Cap-173, step a) (134 mg, 78%) as an orange oil (85% pure) which
was used directly in the next reaction. Rt=1.58 min (Cond.-MD1);
LC/MS: Anal. Calcd. for [M+H]+ C7H10NO2S: 172.05;
found: 172.05.

[0988] Palladium on carbon (10%, 25 mg) was added to a solution of methyl
3-vinylpicolinate (120 mg, 0.74 mmol) in ethanol (10 mL). The suspension
was stirred at room temperature under an atmosphere of hydrogen for 1 h
before it was filtered through Celite and the pad of diatomaceous earth
(Celite®) was washed with methanol. The filtrate was concentrated
down to dryness to yield methyl 3-ethylpicolinate (i.e. Cap-175, step b)
which was taken directly into the next reaction. Rt=1.15 min
(Cond.-MD1); LC/MS: Anal. Calcd. for [M+H]+C9H12NO2:
166.09; found: 166.09.

[1004] An HCV Replicon assay was utilized in the present disclosure, and
was prepared, conducted and validated as described in commonly owned
PCT/US2006/022197 and in O'Boyle et. al. Antimicrob Agents Chemother.
2005 April; 49(4):1346-53. Assay methods incorporating luciferase
reporters have also been used as described (Apath.com).

[1005] HCV-neo replicon cells and replicon cells containing resistance
substitutions in the NS5A region were used to test the currently
described family of compounds. The compounds were determined to have
differing degrees of reduced inhibitory activity on cells containing
mutations vs. the corresponding inhibitory potency against wild-type
cells. Thus, the compounds of the present disclosure can be effective in
inhibiting the function of the HCV NS5A protein and are understood to be
as effective in combinations as previously described in application
PCT/US2006/022197 and commonly owned WO/04014852. It should be understood
that the compounds of the present disclosure can inhibit multiple
genotypes of HCV. Table 2 shows the EC50 (Effective 50% inhibitory
concentration) values of representative compounds of the present
disclosure against the HCV 1b genotype. In one embodiment, compounds of
the present disclosure are inhibitory versus 1a, 1b, 2a, 2b, 3a, 4a, and
5a genotypes. EC50 values against HCV 1b are as follows A (10-350
nM); B (1-9.9 nM); C (0.1-0.99 nM); D (0.0006-0.099 nM).

[1006] The compounds of the present disclosure may inhibit HCV by
mechanisms in addition to or other than NS5A inhibition. In one
embodiment the compounds of the present disclosure inhibit HCV replicon
and in another embodiment the compounds of the present disclosure inhibit
NS5A.

[1007] It will be evident to one skilled in the art that the present
disclosure is not limited to the foregoing illustrative examples, and
that it can be embodied in other specific forms without departing from
the essential attributes thereof. It is therefore desired that the
examples be considered in all respects as illustrative and not
restrictive, reference being made to the appended claims, rather than to
the foregoing examples, and all changes which come within the meaning and
range of equivalency of the claims are therefore intended to be embraced
therein.